Abstract

The increasing use of nanomaterials raises concerns about the long-term effects of chronic nanoparticle exposure on human health. However, nanoparticle exposure is difficult to evaluate non-invasively using current measurement techniques. Here we show that the skin is an important site of nanoparticle accumulation following systemic administration. Mice injected with high doses of gold nanoparticles have visibly blue skin while quantum dot-treated animals fluoresce under ultraviolet excitation. More importantly, elemental analysis of excised skin correlates with the injected dose and nanoparticle accumulation in the liver and spleen. We propose that skin analysis may be a simple strategy to quantify systemic nanoparticle exposure and predict nanoparticle fate in vivo. Our results suggest that in the future, dermal accumulation may also be exploited to trigger the release of ultraviolet and visible light-sensitive therapeutics that are currently impractical in vivo due to limits in optical penetration of tissues at these wavelengths.

(a) Nanoparticles applied topically to the outer surface of the mouse have been shown in previous studies to diffuse through the epidermis to reach the dermal (De) and subcutaneous (Sc) layers of the skin. (b) Systemic administration of nanoparticles by tail-vein injection were found in this study to enter the skin from blood vessels and diffuse into the De and Sc layers of the skin. Our study is focused on the systemic administration and not on the topical application as systemic administration is the most common method for introducing nanoparticle-based drugs and contrast agents into the body.

(a) Mice have a visible blue-purple complexion after tail-vein injection with 6.64 pmol of 15 nm gold nanoparticles functionalized with mPEG after 24 and 504 hours post-injection (HPI). (b) and (c) show the same skin discoloration when injected with 6.64 pmol of 15 nm gold nanoparticles functionalized with transferrin and 100 nm gold nanoparticles functionalized with mPEG respectively. ICP-AES was also used to measure the concentration of the gold nanoparticles. (d) A comparison of gold nanoparticle concentration in small skin biopsies (5 cm2) versus injection dose shows a direct correlation while kinetic plots tracking gold nanoparticle concentration in the skin and lymph nodes (e) demonstrates that nanoparticle clearance from the skin (solid) coincides with an increase in nanoparticle concentration in the axillary lymph nodes (dotted). This suggests that the nanoparticles are cleared from the skin through the lymphatic system. Nanoparticle clearance from the skin levels off at 1.7% dose per gram tissue. (f) Isolated axillary lymph nodes show the gradual accumulation of gold nanoparticles (purple) in the lymph node over time. All error bars denote standard error of the mean values for measurements (n > 3).

(a) Immunohistochemistry stained skin section shows that macrophages (seen in brown) co-localize with nanoparticles (seen in black). Magnified inset clearly shows that nanoparticles can be found in the cytosolic region of macrophages. (b) Microscopy images demonstrate that as injected dose increases, nanoparticles (seen in black) appear to first localize in phagocytic cells (red arrow) then gradually begin spill into the pericellular space of the dermis and finally distribute throughout the dermis (De) and subcutaneous tissue (Sc). Orange dotted lines highlight areas of nanoparticle accumulation. Nanoparticles were not detected in the epidermis (Ep) of the skin. Illustration panel (c–e) shows a pictorial diagram of nanoparticle infiltration into the skin over time. Post-injection, nanoparticles begin to diffuse out of dermal blood vessels (BV) (c) and become taken up by dermal macrophages and dendritic cells (d). Once phagocytic cells become saturated (e) nanoparticles begin to distribute into the pericellular space of the subcutaneous tissue and dermis. Scale bars denote 50 and 200 μm for panels (a) and (b) respectively.

(a & b) Accumulation of gold nanoparticles in the liver and spleen are linearly related to injection dose. (c & d) Accumulation of quantum dots in the liver and spleen are also linearly related to injection dose. (e & f) As the injection dose decreases, we observed a corresponding decrease in the amount of nanoparticles in the liver, spleen, and skin. Interestingly, the quantity of the gold nanoparticles and quantum dots in the liver and spleen is directly related to the skin. This suggests that we can estimate the amount of nanoparticles in the other reticuloendothelial organs by multiplying skin measurements by a numeric factor. For example, the concentration of gold nanoparticles in the liver for an injection dose of 6.64 pmol/g body weight can be obtained by multiplying the measured skin quantity by 2.3. We confirmed that the ratios are statistically similar, using the student’s t-test (p > 0.05). Injection doses for gold nanoparticle and quantum dots were normalized to total surface area to compare between particle types. Error bars denote standard error of the mean values for measurements (n > 3).